Staphylococcus aureus resistente a oxacilina por mecanismos alternativos a la presencia del gen mecA

Staphylococcus aureus is part of skin and mucous membranes colonizing flora, both in humans and animals. However, this opportunistic microorganism is one of the most common and important pathogens isolated both from nosocomial and community environments, responsible for a variety of infections such...

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Autor principal: Weltman, Gabriela Débora
Otros Autores: Di Gregorio, Sabrina
Formato: Tesis de maestría acceptedVersion
Lenguaje:Español
Publicado: Facultad de Farmacia y Bioquímica 2016
Materias:
mecA
Staphylococcus aureus
Oxacilina
Ciencia de la vida
Acceso en línea:http://repositoriouba.sisbi.uba.ar/gsdl/cgi-bin/library.cgi?a=d&c=afamaster&cl=CL1&d=HWA_1631
http://repositoriouba.sisbi.uba.ar/gsdl/collect/afamaster/index/assoc/HWA_1631.dir/1631.PDF
Aporte de:
Repositorio Digital de la Universidad de Buenos Aires (UBA) de Universidad de Buenos Aires
id I28-R145-HWA_1631
record_format dspace
institution Universidad de Buenos Aires
institution_str I-28
repository_str R-145
collection Repositorio Digital de la Universidad de Buenos Aires (UBA)
language Español
orig_language_str_mv spa
topic mecA
Staphylococcus aureus
Oxacilina
Ciencia de la vida
spellingShingle mecA
Staphylococcus aureus
Oxacilina
Ciencia de la vida
Weltman, Gabriela Débora
Staphylococcus aureus resistente a oxacilina por mecanismos alternativos a la presencia del gen mecA
topic_facet mecA
Staphylococcus aureus
Oxacilina
Ciencia de la vida
description Staphylococcus aureus is part of skin and mucous membranes colonizing flora, both in humans and animals. However, this opportunistic microorganism is one of the most common and important pathogens isolated both from nosocomial and community environments, responsible for a variety of infections such as endocarditis, osteomyelitis, and sepsis.\nFour clinical isolates of S. aureus recovered from a patient who suffered four consecutive episodes of bacteremia were characterized in this work. The first three isolates were susceptible to cefoxitin (FOX) and oxacillin (OXA), while the latter was resistant to OXA.\nThe isogenicity of the four isolates was determined by PFGE. Genotyping indicated that these isolates belong to ST8, spa type 024. This lineage is not prevalent in Argentina. The first isolate (SAMS1) and the last one (SA2) were further studied, considering the rare phenotype observed in SA2 (FOX sensitivity and OXA resistance).\nMolecular detection of the mecA gene, the most common mechanism associated with OXA resistance, was negative. The presence of mecC gene and recombinase genes described in different SCCmec were also discarded. Consequently, alternative mechanisms that could explain this phenotype were evaluated.\nNo mutations in pbp4 gene were detected among isolates.\nThe SA2 isolate showed higher ?-lactamase activity than the SAMS1 isolate. However, no mutations in the bla operon, that could justify this overproduction, were found.\nComparison of SA2 and SAMS1 pbp2 gene sequences revealed a mutation at nucleotide 1350, resulting in an Ala450Asp substitution. This change was not detected in the SAMS3 strain (isolated in the third episode of bacteremia), suggesting that this mutation could be linked with the emergence of oxacillin resistance in SA2\nA tridimensional model showed the replacement of a hydrophobic aminoacid such\nas alanine, by an acid one as aspartic, placed in a hydrophobic environment close to the\nactive site cavity of the transpeptidase domain of SA2 PBP2. This change would generate\ndistortions that could hinder the OXA molecule entrance.\nBoth ?-lactamase overproduction and PBP2 Ala450Asp substitution described in\nSA2 strain, would contribute to the resistance phenotype described herein. The possible\ncoexistence of other molecular mechanisms that could contribute to the oxacillin\nresistance could not be discarded.\nDetection of oxacillin resistant, mec negative isolates, represents a major challenge\nfor clinical laboratories, since they are not detected using FOX disc diffusion method.\nThis work demonstrates the emergence of OXA resistant S. aureus isolates in Argentina\nthat are not detected with current CLSI recommendations.
author2 Di Gregorio, Sabrina
author_facet Di Gregorio, Sabrina
Weltman, Gabriela Débora
format Tesis de maestría
Tesis de maestría
acceptedVersion
author Weltman, Gabriela Débora
author_sort Weltman, Gabriela Débora
title Staphylococcus aureus resistente a oxacilina por mecanismos alternativos a la presencia del gen mecA
title_short Staphylococcus aureus resistente a oxacilina por mecanismos alternativos a la presencia del gen mecA
title_full Staphylococcus aureus resistente a oxacilina por mecanismos alternativos a la presencia del gen mecA
title_fullStr Staphylococcus aureus resistente a oxacilina por mecanismos alternativos a la presencia del gen mecA
title_full_unstemmed Staphylococcus aureus resistente a oxacilina por mecanismos alternativos a la presencia del gen mecA
title_sort staphylococcus aureus resistente a oxacilina por mecanismos alternativos a la presencia del gen meca
publisher Facultad de Farmacia y Bioquímica
publishDate 2016
url http://repositoriouba.sisbi.uba.ar/gsdl/cgi-bin/library.cgi?a=d&c=afamaster&cl=CL1&d=HWA_1631
http://repositoriouba.sisbi.uba.ar/gsdl/collect/afamaster/index/assoc/HWA_1631.dir/1631.PDF
work_keys_str_mv AT weltmangabrieladebora staphylococcusaureusresistenteaoxacilinapormecanismosalternativosalapresenciadelgenmeca
_version_ 1766017480631779328
spelling I28-R145-HWA_16312019-09-27 Staphylococcus aureus is part of skin and mucous membranes colonizing flora, both in humans and animals. However, this opportunistic microorganism is one of the most common and important pathogens isolated both from nosocomial and community environments, responsible for a variety of infections such as endocarditis, osteomyelitis, and sepsis.\nFour clinical isolates of S. aureus recovered from a patient who suffered four consecutive episodes of bacteremia were characterized in this work. The first three isolates were susceptible to cefoxitin (FOX) and oxacillin (OXA), while the latter was resistant to OXA.\nThe isogenicity of the four isolates was determined by PFGE. Genotyping indicated that these isolates belong to ST8, spa type 024. This lineage is not prevalent in Argentina. The first isolate (SAMS1) and the last one (SA2) were further studied, considering the rare phenotype observed in SA2 (FOX sensitivity and OXA resistance).\nMolecular detection of the mecA gene, the most common mechanism associated with OXA resistance, was negative. The presence of mecC gene and recombinase genes described in different SCCmec were also discarded. Consequently, alternative mechanisms that could explain this phenotype were evaluated.\nNo mutations in pbp4 gene were detected among isolates.\nThe SA2 isolate showed higher ?-lactamase activity than the SAMS1 isolate. However, no mutations in the bla operon, that could justify this overproduction, were found.\nComparison of SA2 and SAMS1 pbp2 gene sequences revealed a mutation at nucleotide 1350, resulting in an Ala450Asp substitution. This change was not detected in the SAMS3 strain (isolated in the third episode of bacteremia), suggesting that this mutation could be linked with the emergence of oxacillin resistance in SA2\nA tridimensional model showed the replacement of a hydrophobic aminoacid such\nas alanine, by an acid one as aspartic, placed in a hydrophobic environment close to the\nactive site cavity of the transpeptidase domain of SA2 PBP2. This change would generate\ndistortions that could hinder the OXA molecule entrance.\nBoth ?-lactamase overproduction and PBP2 Ala450Asp substitution described in\nSA2 strain, would contribute to the resistance phenotype described herein. The possible\ncoexistence of other molecular mechanisms that could contribute to the oxacillin\nresistance could not be discarded.\nDetection of oxacillin resistant, mec negative isolates, represents a major challenge\nfor clinical laboratories, since they are not detected using FOX disc diffusion method.\nThis work demonstrates the emergence of OXA resistant S. aureus isolates in Argentina\nthat are not detected with current CLSI recommendations. Fil: Weltman, Gabriela Débora. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Buenos Aires, Argentina Di Gregorio, Sabrina Facultad de Farmacia y Bioquímica Mollerach, Marta Weltman, Gabriela Débora 2016-09-19 Staphylococcus aureus, forma parte de la microbiota colonizante de piel y mucosas en humanos y animales. Sin embargo, este patógeno oportunista es uno de los patógenos más frecuentes e importantes, aislado tanto en el ámbito nosocomial como en la comunidad, responsable de una gran variedad de infecciones tales como endocarditis, osteomielitis y sepsis.\nEn el presente trabajo se caracterizaron cuatro aislamientos clínicos de S. aureus recuperados de un paciente que padeció cuatro episodios sucesivos de bacteriemia. Los primeros 3 aislamientos fueron sensibles a cefoxitina (FOX) y oxacilina (OXA), mientras que el último resultó resistente a OXA.\nSe determinó la isogenicidad de los cuatro aislamientos recuperados del paciente por la técnica de PFGE. La genotipificación indicó que estos aislamientos pertenecen al ST8, y spa tipo t024, linaje que no se encuentra dentro de los más frecuentes en Argentina. Se profundizó el estudio del primer aislamiento, SAMS1, y del último, SA2 dado su fenotipo poco frecuente en nuestro medio (sensibilidad a FOX y resistencia a OXA)\nLa detección molecular del gen mecA que codifica la PBP2a, mecanismo más frecuente asociado a resistencia a oxacilina, dio resultado negativo. Se descartó también la presencia del gen mecC y de las diferentes recombinasas descriptas en los SCCmec. En consecuencia, se evaluaron mecanismos alternativos que pudiesen explicar este fenotipo.\nNo se detectaron mutaciones en el gen pbp4 entre los aislamientos.\nEl aislamiento SA2 presentó mayor actividad de ?-lactamasas que el aislamiento SAMS1. Sin embargo, no se encontraron mutaciones en el operón bla, que pudieran justificar esta hiperproducción.\nSe pudo evidenciar también, la presencia de una mutación en la posición 1350 del\ngen pbp2 de la cepa SA2 respecto de la SAMS1, que resultó en una sustitución\nAla450Asp. Este cambio no se evidenció en la cepa SAMS3 (aislada en el tercer episodio\nde bacteriemia), lo que sugiere que esta mutación se podría vincular con la emergencia\nde resistencia a oxacilina en SA2.\nEl análisis del efecto de esta mutación con modelos tridimensionales muestra el\nreemplazo de un aminoácido hidrofóbico como la alanina, por uno ácido como el\naspártico, en un entorno hidrofóbico y cercano a la cavidad del sitio activo del dominio\ntranspeptidasa de la PBP2 de SA2. La carga introducida en ese entorno hidrofóbico\nproduciría una distorsión del ?-hélice y del cercano bolsillo de reacción, dificultando así\nel ingreso de la molécula de OXA.\nPosiblemente, tanto la hiperproducción de ?-lactamasa como la mutación que\nresulta en la sustitución Ala450Asp en PBP2, descriptas en la cepa SA2, contribuyan al\nfenotipo de resistencia observado. No se descarta la posible coexistencia de otros\nmecanismos moleculares que contribuyan a la resistencia a este antibiótico.\nLa detección de aislamientos resistentes a oxacilina pero carentes del gen mec,\nrepresenta un gran desafío para los laboratorios clínicos, dado que no se detectan\nutilizando el disco de FOX. Si bien el CLSI recomienda, dentro de las pruebas cualitativas,\nel uso del disco de FOX para informar la sensibilidad a antibióticos ?-lactámicos, este\ntrabajo demuestra la emergencia en Argentina de aislamientos de S. aureus resistentes\na OXA que no se detectan con ese método y serían reportados como SAMS. application/pdf Centrón, Daniela Gutkind, Gabriel Basualdo Farjat, Juan mecA Staphylococcus aureus Oxacilina spa Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica info:eu-repo/semantics/openAccess http://creativecommons.org/licenses/by-nc-nd/2.5/ar/ Ciencia de la vida Staphylococcus aureus resistente a oxacilina por mecanismos alternativos a la presencia del gen mecA info:eu-repo/semantics/masterThesis info:ar-repo/semantics/tesis de maestría info:eu-repo/semantics/acceptedVersion http://repositoriouba.sisbi.uba.ar/gsdl/cgi-bin/library.cgi?a=d&c=afamaster&cl=CL1&d=HWA_1631 http://repositoriouba.sisbi.uba.ar/gsdl/collect/afamaster/index/assoc/HWA_1631.dir/1631.PDF

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